Endogenous Indole-3-acetic Acid Levels Research Articles

Effects of Exogenous BAP, GA3, and ABA on Endogenous Auxin and Rooting of Grapevine Hardwood Cuttings.

The effects of applications of 6-benzylaminopurine (BAP), gibberellic acid (GA3), and abscisic acid (ABA) on the root formation of grapevine hardwood cuttings were investigated. BAP suppressed rooting of grapevine cuttings completely, whereas GA3 and ABA inhibited it temporarily. Endogenous indoleacetic acid (IAA) in cuttings treated with BAP increased linearly to 12.6 and 19.4 times per cutting and per g fresh weight, respectively, over the control for 30 days after planting. In cuttings treated with GA3, the level of diffusible IAA was higher than that of control cuttings through the experimental period; the fluctuation pattern of extractable IAA was similar to that of the control. In cuttings treated with ABA, the extractable IAA content was significantly lower than that in control cuttings 10 days after planting. The fluctuation pattern of endogenous IAA in BAP treatment was different from that in GA3 and ABA treatments. Cuttings treated with BAP presumably failed to root because the level of endogenous IAA was unbalanced. GA3 may inhibit root formation directly whereas ABA may cause inhibition of rooting by depressing the bud activity.

Endogenous indole-3-acetic acid during adventitious root formation inPopulus £canadensis Moench.

Indole-3-butyric acid (IBA), phenylacetic acid (PAA) and naphthaleneacetic acid (NAA) were applied at a concentration of 10-4 mol dm-3 to stem cutting bases ofPopulus x canadensis Moench. During adventitious root formation, the content of indole-3-acetic acid (IAA) in cutting bases was estimated using the fluorimetric method. In the control variant, a rapid increase in endogenous IAA appeared after 24-h cultivation followed by gradual decrease during the following days. In contrast, the variants treated with IBA, PAA, and especially NAA exhibited firstly a decrease in endogenous IAA content and only afterwards an increase, reaching a maximum 48 h after excision. As root regeneration proceeded gradually, a decrease in the level of endogenous IAA occurred in all treatments. The first adventitious roots appeared in all treatments after 216-h cultivation.

Decarboxylation of lndole‐3‐Acetic Acid and Inhibition of Growth in Avena sativa Seedlings by Plant‐Derived Photosensitizers*

Abstract— A number of plant phototoxins, when supplemented with UVA (320–400 nm) radiation, are capable of sensitizing the decomposition of indole‐3‐acetic acid (IAA), as measured by release of 14CO2 from carboxyl‐labeled IAA. Alpha‐terthienyl (αT) and harmine caused significant rates of IAA decarboxylation at concentrations as low as 1 nM and were approximately 80% as effective as riboflavin and flavin mononucleotide. Partial inhibition by sodium azide indicates that the aT‐induced decarboxylation of IAA is predominately, but not entirely, a type II reaction mediated by singlet oxygen. Based on changes in UV ab sorption spectra, it appears that the hormones gibberellic acid, abscisic acid and 6‐benzylaminopurine (a cytokinin) are less susceptible to photosensitized decomposition than is IAA. Alpha‐terthienyl plus UVA also inhibited elongation growth and reduced endogenous IAA levels in Avena sativa L. coleoptile sections and promoted senescence in intact Avena seedlings. These results confirm the allelopathic potential of plant photosensitizers such as αT and indicate that the phytohormone IAA may represent an additional target for the action of photosensitizers.

The response to auxin of rapeseed (Brassica napus L.) roots displaying reduced gravitropism due to transformation by Agrobacterium rhizogenes.

It has recently been documented that, compared to untransformed controls, the roots of oilseed rape (Brassica napus L. CV CrGC5) seedlings transformed by Agrobacterium rhizogenes A4 show a reduced gravitropic reaction (Legue et al. 1994, Physiol Plant 91: 559-566). After stimulation at 90 degrees C or 135 degrees, the transformed root tips curve. but never reach a vertical orientation. In the present study, we investigated the causes of reduced gravitropic bending observed in stimulated transformed root tips. First, we localized the gravitropic curvature in normal and in transformed roots after 1.5 h of stimulation. The cells involved in root curvature (target cells) corresponded at the cellular level to the apical part of the zone of increasing cell length. In transformed roots grown in the vertical position, these cells showed a reduction in cell length compared to controls. Because auxin is considered to be the gravitropic mediator, the response of normal and transformed roots to exogenous auxin was studied. Indole-3-acetic acid (IAA) was applied along the first 3 mm using resin beads loaded with the hormone. In comparison to normal roots, transformed roots showed reduced bending toward the bead at all points of bead application. Moreover, the cells which responded to IAA corresponded to the target cells involved in the gravitropic reaction. The level of endogenous IAA was lower in transformed roots. Thus, it was concluded that the modified behavior of transformed roots during gravitropic stimulation could be due to differences either in IAA levels or in reactivity of the target cells to the message from the cap.

Paclobutrazol affects the extension growth and the levels of endogenous IAA of almond seedlings

The effect of paclobutrazol on plant height and the concentration of free indole-3-acetic acid (IAA) in almond seedlings grown in 0.5 L pots with moist perlite and subjected to 15°C and photoperiod of 16 hours was studied.

Changes in ABA, IAA and GAs Contents in Reproductive Organs of Satsuma Mandarin.

The endogenous levels of abscisic acid (ABA), indole-3-acetic acid (IAA) and gibberellins (GAs) in reproductive organs of satsuma mandarin (Citrus unshiu Marc. cv. Ozaki) were determined by gas chromatograph (GC)-electron capture detector, GC-mass spectrometry-selective ion monitoring and dwarf rice bioassay. Their probable roles are discussed.1. As anthesis approached, IAA concentration in styles and stamens did not change, while ABA concentration of styles increased six-fold. Until 39 days after flowering (DAF), the fresh weigth of fruitlets increased dramatically, while that of 'sepals' which included floral disks, sepals and receptacles increased a little.2. Fruitlets attained their maximum concentration of ABA 4 DAF and then declined rapidly. Its level remained much lower than that of 'sepals' from 7 DAF. IAA concentrations of fruitlets reached a maximum 7 DAF, and thereafter decrease; that in the 'sepals' increased from anthesis to 31 DAF surpassing that of fruitlets about 15 DAF.3. GAs concentrations of fruitlets continued to increase from 7 to 31 DAF, and decreased sharply 39 DAF.4. Because there was a sequence of peaks of endogenous ABA, IAA and GAs levels in fruitlets, it seemed that these hormones may play a sequential and synergistic role in the retention and growth of fruitlets.

Assimilation of exogenous 2′-14C-indole-3-acetic acid and 3′-14C-tryptophan exposed to the roots of three wheat varieties

This study was conducted to determine if plants can assimilate indole-3-acetic acid (IAA) from rooting media and if exogenous L-tryptophan (L-TRP) can be assimilated and converted by plants into auxins. The addition of 2′-14C-IAA (3.7 kBq plant-1) to wheat (Triticum aestivum L.) seedlings of three varieties grown in nutrient solution resulted in the uptake (avg.=7.6%) of labelled IAA. Most of the label IAA was recovered in the shoot (avg.=7.2%) with little accumulation in the root (avg.=0.43%). A portion of the assimilated IAA-label in the plant was identified by co-chromatography and UV spectral confirmation as IAA-glycine and IAA-aspartic acid conjugates. Little of the assimilated IAA label was found as free IAA in the wheat plants. These same assimilation patterns were observed when 2′-14C-IAA was added to wheat plants grown in sterile and nonsterile soil. In contrast, the wheat varieties assimilated considerably less (avg.=1.3%) of the added microbial IAA precursor, 3′-14C-L-TRP (3.7 kBq plant-1) and thus much lower amounts of IAA conjugates were detected. Glasshouse soil experiments revealed that 2 out of 3 wheat varieties had increased growth rates and increased yields when L-TRP (10-5 and 10-7M) was added to the root zone. It is surmised that this positive response is a result of microbial auxin production within the rhizosphere upon the addition of the precursor, L-TRP. The amino acid composition of the root exudates plays a critical role in microbial production of auxins in the rhizosphere. This study showed that wheat roots can assimilate IAA from their rooting media, which will supplement the endogenous IAA levels in the shoot tissue and may positively influence plant growth and subsequent yield.

Growth patterns and endogenous indole‐3‐acetic acid concentrations in current‐year coppice shoots and seedlings of two Betula species

Apical dominance, internode elongation, radial growth and xylem cell size in coppice and apical shoots of Betula pubescens B. Pendula were determined and related to endogenous indole‐3‐acetic acid (IAA) levels, measured by gas chromatography‐selected ion monitoring in the apical bud and at three positions along the stem. The effects of defoliation and debudding on morphological and anatomical characters and endogenous IAA levels were also investigated. The coppice shoots displayed superior stem elongation and increased branching during the initial phase of growth, after which their growth pattern was similar to that of the seedlings; however, their radial growth was greater throughout the experiment. Both plant types produced smaller‐sized xylem cells at the top of the shoot than at the bottom with coppice shoots tending to form larger tracheids and smaller vessels than the seedlings. There was no consistent difference in IAA concentration between the coppice shoots and the seedlings. Defoliation and debudding reduced the IAA level in the stem within 36 h and it was still low after 25 days. Although the extent of the IAA decrease was similar in both coppice shoots and seedlings, the treatments affected the morphological and anatomical characters differently in the two plant types. The results suggest that the observed differences between seedlings and coppice shoots were not mediated through a drastic change in IAA level.

Involvement of Abscisic Acid in Ethylene-Induced Cotyledon Abscission in Cotton Seedlings.

Cotton (Gossypium hirsutum L. cv LG102) seedlings raised from seeds exposed to 100 [mu]M norflurazon (NFZ) during imbibition contained reduced levels of free abscisic acid (ABA) and were visibly achlorophyllous. Exposure of untreated cotton seedlings to ethylene concentrations >1 [mu]L/L for 24 h resulted in cotyledon abscission. In contrast, exposure of NFZ-treated seedlings to concentrations of ethylene [less than or equal to]50 [mu]L/L elicited no cotyledon abscission. Application of ABA, an ABA analog, or jasmonic acid to NFZ-treated seedlings restored ethylene-induced abscission. Isolated cotyledonary node explants prepared from NFZ-treated seedlings exhibited an altered dose-response pattern of ethylene-induced petiole abscission. Endogenous levels of free IAA were unaltered in NFZ-treated seedlings. Ethylene treatment (50 [mu]L/L, 24 h) had no effect on free indoleacetic acid (IAA) levels in either control or NFZ-treated seedlings. Levels of conjugated (ester plus amide) IAA were substantially increased in NFZ-treated seedlings regardless of ethylene treatment. These results indicate that endogenous ABA plays an essential, but physiologically undefined, role in ethylene-induced cotyledon abscission in cotton.

Endogenous levels of IAA in relation to parthenocarpy in cucumber ( Cucumis sativus L.)

The role of endogenous indole-3-acetic acid (IAA) on parthenocarpic fruit set in cucumber was investigated. Parthenocarpic fruit set was closely related to the accumulation of IAA in the ovary. Application of N-1-napthylphthylamic acid (naptalam) and 2,3,5-triiodobenzoic acid (TIBA), auxin transport inhibitors, to the ovary or peduncle significantly increased the IAA content in the ovary. Exogenously applied auxins, naphthaleneacetic acid (NAA) and 4-chlorophenoxyacetic acid (4-CPA), gibberellic acid (GA 3) and cytokinins, N-(2-chloro-4-pyridyl)-N′-phenylurea (CPPU) and 6-benzylaminopurine (BA), also increased it. It is suggested that endogenous IAA is the main hormonal factor for parthenocarpic fruit set and exogenously applied hormones are considered to promote IAA accumulation in the ovary.

Wounding Nicotiana tabacum Leaves Causes a Decline in Endogenous Indole-3-Acetic Acid

We have previously observed that auxin can act as a repressor of the wound-inducible activation of a chimeric potato proteinase inhibitor II-CAT chimeric gene (pin2-CAT) in transgenic tobacco (Nicotiana tobacum) callus and in whole plants. Therefore, this study was designed to examine endogenous levels of indole-3-acetic acid (IAA) in plant tissues both before and after wounding. Endogenous IAA was measured in whole plant tissues by gas chromatography-mass spectrometry using an isotope dilution technique. (13)C-Labeled IAA was used as an internal standard. The endogenous levels of IAA declined two- to threefold within 6 hours after a wound. The kinetics of auxin decline are consistent with the kinetics of activation of the pin2-CAT construction in the foliage of transgenic tobacco.

Tracheid production in response to indole-3-acetic acid varies with internode age in Pinus sylvestris stems

Different concentrations of indole-3-acetic acid (IAA) in lanolin were applied to the cambial region of approximately 10- and 34-year-old internodes in the main stem of Pinus sylvestris (L.) trees during the tracheid production period. After 5 weeks of treatment, the radial width of xylem produced in both ages of internode was positively related to exogenous IAA concentration measured at 0, 1 and 3 cm directly below the application site. Tracheid production in response to exogenous IAA in the 34-year-old internode was approximately one-half of that in the 10-year-old internode. The endogenous IAA level in the 7-, 17- and approximately 34-year-old internodes of similar trees was measured by radioimmunoassay, using gas chromatography-selected ion monitoring-mass spectrometry for validation. No consistent relationship was found between xylem radial width and IAA concentration. The data indicate that the cambium's ability to respond to exogenous IAA is qualitatively the same in 1-year-old shoots and older internodes. However, as the internode ages, there is a decrease in the extent of the response and in the optimal IAA level for inducing tracheid production.

Level of endogenous indole‐3‐acetic acid in the stem of Pinus sylvestris in relation to the seasonal variation of cambial activity

Abstract. Gas chromatography – selected ion monitoring – mass spectrometry was used to measure the level of indole‐3‐acetic acid (IAA) in the cambial region at the top and bottom of the branchless portion of the main stem of three large Scots pine trees, at weekly intervals from 28 April to 13 July. During this period, the cambium reactivated from the dormant state and entered its ‘grand’ period of xylem and phloem production, which was monitored by microscopy. The total amount of IAA (ng cm−2) increased steadily from 28 April until late June, and thereafter remained constant. In contrast, the concentration of IAA (ng g−1 fresh weight) was high at the start of cambial reactivation, declined when the number of differentiating tracheids began to increase, and then rose as the number of cells decreased. The timing and magnitude of the changes in xylem and phloem production and in IAA level were similar at the two sampling positions. It is concluded that the seasonal changes in cambial activity in the conifer stem cannot be ascribed simply to a fluctuation in the level of endogenous IAA in the cambial region.

Tracheid Production in Response to Changes in the Internal Level of Indole-3-Acetic Acid in 1-Year-Old Shoots of Scots Pine

Different concentrations of indole-3-acetic acid (IAA) were applied in lanolin to 1-year-old shoots of Pinus sylvestris (L.) in a manner known to stimulate cambial activity. The internal concentration of free IAA was measured at a distance below the application point by combined gas chromatography-selected ion monitoring-mass spectrometry using [(13)C(6)]IAA as a quantitative internal standard, and related to the production of tracheids at the same site. The experiment was performed with: (a) debudded cuttings, where the major source of endogenous IAA, the apical buds, were replaced with exogenous IAA, and (b) intact, attached shoots, where endogenous IAA was supplemented by applying IAA around the circumference of the shoot. In both experimental systems, an increase in the internal IAA level was positively related to increased tracheid production. It was also demonstrated that the concentration of internal IAA measured at the sampling site was comparable with endogenous IAA levels found in intact control shoots, and that a wide range of applied IAA concentrations was associated with a relatively small range of internal IAA levels.

Changes in the concentration of indole‐3‐acetic acid during the growth of etiolated lupin hypocotyls

The longitudinal distribution of unaltered radioactive indole‐3‐acetic acid (IAA), after application of [5‐3H]‐IAA to decapitated etiolated lupin hypocotyls. exhibited a wave‐like pattern similar to that obtained with endogenous IAA. Waves of radioactive IAA were localizated both in the elongation zone and in the non‐growing basal region of the hypocotyl. These IAA waves were transient because of basipetal polar transport and metabolism of IAA.The level of endogenous IAA in different zones of the hypocotyl varied with age, following a wave‐like pattern. During the elongation period of each zone, IAA was parallel to the bell‐shaped curve of the growth rate. In addition, a role in secondary cell wall deposition is suggested for the other IAA wave that appeared after the cell elongation period, since an electron microscopic morphometric analysis of the cell wall showed that the cell wall thickness increased once the cell elongation ceased.As the oscillation of endogenous IAA level occured in both space (distribution along the hypocotyl) and time (variation with age), it is suggested that the level of IAA really depended on the growth status of the cells. The response of the cells to the positional information submitted by the auxin waves as regards the growth status of the cell is discussed.

Ethylene-Enhanced Catabolism of [14C]Indole-3-Acetic Acid to Indole-3-Carboxylic Acid in Citrus Leaf Tissues

Exogenous [(14)C]indole-3-acetic acid (IAA) is conjugated in citrus (Citrus sinensis) leaf tissues to one major substance which has been identified as indole-3-acetylaspartic acid (IAAsp). Ethylene pretreatment enhanced the catabolism of [(14)C]IAA to indole-3-carboxylic acid (ICA), which accumulated as glucose esters (ICGIu). Increased formation of ICGIu by ethylene was accompanied by a concomitant decrease in IAAsp formation. IAAsp and ICGIu were identified by combined gas chromatography-mass spectrometry. Formation of ICGIu was dependent on the concentration of ethylene and the duration of the ethylene pretreatment. It is suggested that the catabolism of IAA to ICA may be one of the mechanisms by which ethylene reduces endogenous IAA levels.

Cytokinin-to-Auxin Ratios and Morphology of Shoots and Tissues Transformed by a Chimeric Isopentenyl Transferase Gene

Tissues transformed with the isopentenyl transferase (ipt) gene cloned from the T-DNA region of the Ti plasmid or with the ipt gene placed under the control of the cauliflower mosaic virus 35S promoter (35S-ipt) were analyzed for auxin and cytokinin. Free and total indole-3-acetic acid (IAA) levels in 35S-ipt-transformed Nicotiana tabacum and cucumber cells were reduced by 12 to 78% in comparison to untransformed tissues. In contrast, free IAA concentrations in 35S-ipt-transformed Nicotiana plumbaginifolia were almost three times those of untransformed tissues, while total IAA levels were not significantly affected. Cytokinin levels in these antibodies were elevated an average of 300-fold resulting in a 24- to over 2000-fold increase in the cytokinin-to-auxin ratios. High cytokinin-to-auxin ratios correlated with the shooty phenotype of transformed tissues propagated in vitro in the absence of added growth hormones. We conclude that increased endogenous cytokinin-to-auxin ratios can induce cells to undergo morphogenesis and that elevated cytokinin levels can also induce auxin-autonomous growth of transformed tissues without causing a corresponding increase in endogenous IAA levels.

Fluctuation of the endogenous IAA level in rice during its life cycle.

Endogenous auxins in the shoots and ears of rice were investigated. Indole-3-acetic acid (IAA) and indole-3-carboxylic acid (ICA) was identified by GC-MS, the endogenous level of IAA being much higher than that of ICA. To analyze the fluctuation of endogenous IAA level throughout the life cycle of rice, a rapid and effective procedure, using HPLC and a fluorescence detector, was developed. The level of IAA in shoots was 10 —26ng/g fr. wt., while that in ears was 10 to 100 times higher. The level of IAA conjugate in ears was also much higher than that in shoots. These results show that the biosynthesis of IAA occurs at an early stage of seed development, and it is also suggested that IAA may play a role in regulating the reproductive growth of rice.

Levels of Indole-3-Acetic Acid in Lemna gibba G-3 and in a Large Lemna Mutant Regenerated from Tissue Culture

Large changes in indole-3-acetic acid (IAA) levels occur during growth of Lemna gibba G-3 in sterile culture. The levels of IAA were measured in plants during a 45 day growth cycle using HPLC and isotope dilution analysis followed by selected ion current monitoring GC-MS analysis with (13)C(6)-IAA as the internal standard. Even though the rate of plant growth remained constant over the entire growth period, IAA levels ranged from a high of 222 to a low of 6 nanograms per gram fresh weight. A Lemna mutant (jsR(1)) which has a giant phenotype was obtained by regeneration from primary callus cultures. Microspectrofluorometry of diamidino-2-phenylindole stained cells showed that jsR(1) has the same amount of DNA per nucleus as the parent line (PL). All jsR(1) cell types measured are about 1.5 times larger than in PL. The endogenous levels of IAA per gram fresh weight were higher in jsR(1) at several stages of the plant culture cycle as compared to PL. This difference ranged from 1.2 to over 100 times as much. While PL showed only one high peak at day 9, jsR(1) had IAA levels of 480 and 680 nanograms per gram fresh weight at days 9 and 45, respectively. Throughout the midculture stage of the growth cycle (20-28 days) both jsR(1) and PL had IAA levels in the range of 9 to 14 nanograms per gram fresh weight. In contrast to PL, at day 45, jsR(1) had no detectable ester or amide conjugates of IAA. These changes in IAA levels were determined in sterile plant cultures and thus cannot be attributed to bacterial or fungal activity.

Endogenous Abscisic Acid and Indole-3-Acetic Acid and Somatic Embryogenesis in Cultured Leaf Explants of Pennisetum purpureum Schum.

Effects of application in vivo of glyphosate, fluridone, and paclobutrazol to glasshouse-grown donor plants of Pennisetum purpureum Schum. on endogenous levels of abscisic acid (ABA) and indole-3-acetic acid (IAA) in young leaves and on somatic embryogenesis in cultured leaf explants were studied. Treatment of plants with glyphosate (100 milligrams per liter) resulted in elevated levels of endogenous ABA and IAA in young leaves. In contrast, paclobutrazol (50% active ingredient; 200 milligrams per liter) did not alter the endogenous levels of ABA and IAA. Fluridone (100 milligrams per liter) markedly inhibited synthesis of ABA and leaf explants from fluridone-treated plants lost the capacity for somatic embryogenesis. Explants from glyphosate- or paclobutrazol-treated plants did not show any reduction in embryogenic capacity when compared with untreated control plants. Glyphosate and fluridone were also incorporated into the culture media at various concentrations (0 to 20 milligrams per liter) to study their effects in vitro on somatic embryogenesis in leaf explants from untreated, field-grown plants. Glyphosate was inhibitory to somatic embryogenesis but only at concentrations above 5 milligrams per liter. Fluridone inhibited somatic embryogenesis at all concentrations tested. Inhibition of somatic embryogenesis by fluridone, by either in vivo or in vitro application, could be overcome partially by (+/-)-ABA added to the culture medium. Exogenous application of (+/-)-ABA enhanced somatic embryogenesis and reduced the formation of nonembryogenic callus. Application of IAA or gibberellic acid (GA(3); >5 milligrams per liter) was inhibitory to somatic embryogenesis. These results indicate that endogenous ABA is one of the important factors controlling the embryogenic capacity of leaf explants in Napier grass.